Featured image and figures
used with permission via ACS Editors’ Choice open access

When we think of medicine, our first thought is often about
small molecules like the antibiotic penicillin or the analgesic acetaminophen.
Proteins, however, can also be used to treat human disease such as when the peptide
hormone insulin is used to treat diabetes. Since many of the jobs inside cells
are completed by proteins, it could be very beneficial to be able to deliver
specific protein drugs directly into cells, such as when certain cells don’t
make as much of a protein as they should. Proteins can be fairly large
molecules though, so it can be difficult for them to move from the blood stream
into cells by crossing the cell membranes that are intended to keep foreign
substances out. For that reason, scientists are developing new ways to trick
cells into engulfing proteins, much like a dog’s medicine might be hidden
inside food.

The basis for this recent strategy is to encompass many
proteins—in this case, insulin—within the framework of many oblong nanoparticles
(NPs). These nanoparticles are small and stable, so they can travel throughout
the bloodstream, and they are porous so they can carry cargo like proteins
within their pores. These NPs, like many in recent years, are made of
Metal-Organic Frameworks (MOFs), meaning a combination of organic and metal (in
this case, Zirconium) building blocks. This combination gives peptides like
insulin a lot of sticky chemical groups to hold onto, making this a good delivery
system. In order to find the best insulin-delivering NP, they tried using two
different MOFs: one called NU-1000 (using the organic molecule H4-TBAPy,
Fig. 1B) and the other called PCN-222 (using another organic molecule, H4-TCPP-H2,
Fig. 1B).

However, these MOF NPs tend to aggregate (clump together)
which hinders them from flowing freely through body fluids to reach their
targets. They are also highly positively charged which makes it difficult for
them to enter cells and can even cause death in the cells they interact with.
Thus, the scientists Wang and colleagues recognized the need for a coating
around these nanoparticles to combat these downsides. Luckily, there is a very
common and easy to work with biomolecule with a net-negative charge: DNA.

DNA is made up of nucleotide bases attached to each other via a backbone of deoxyribose sugars and
negatively charged phosphate groups (Fig. 1C). The negatively charged phosphate
group can interact with the positively charged zirconium metal in the MOF NP.
This leads to a hair-like coating of DNA strands surrounding the NP. To
determine whether the insulin and DNA would both cling to the MOF NPs, they
labeled insulin with a green fluorescent dye and DNA with a red fluorescent dye
and looked at their MOFs using a confocal microscope. Indeed, they did see the
DNA and insulin (Fig. 2A) in the same oblong shapes as the MOFs (Fig. 2C,D). Now
that the NPs had been fully assembled, they need to be tested for their ability
to remain in suspension (i.e. not aggregate), enter cells, and not cause cell
death.

After determining
that the DNA-MOF-NPs did not have their predecessors’ aggregation problems, the
scientists tested whether they could enter human ovarian cancer cells better
than free insulin and free DNA could. In this experiment, they again labeled insulin
with green fluorescent dye and DNA with red fluorescent dye and looked at their
location within cells through confocal microscopy (Fig. 4A-C, right). In
samples with only free insulin/DNA, there wasn’t much sign of either inside the
cells, but the cells with either DNA-MOF-NP showed fluorescence from both DNA
and insulin. Another method—flow cytometry—was used to quantify the amount of
green and red fluorescence overlapping in many cells. This can be seen on the
graphs as a heat map of fluorescence intensity, and the more of the heat map
that lies in the upper-right quadrant of the graph, the more DNA-MOF-NP uptake there
was (Fig. 4A-C, left). The scientists also tested the number of living and dead
cells before and after DNA-MOF-NP treatment, and found no change for either
DNA-NU-1000 or DNA-PCN-222 up to 1nM concentration.

These DNA-coated Molecular Organic Framework Nanoparticles
show a lot of promise as a potential delivery system to get protein drugs into
human cells, and thus there is a lot of work still to be done. How well can
other proteins besides insulin be transported? What are the potential
side-effects, whether from the whole DNA-MOF-NPs or their degradation products?
As more years of research find answers to these questions, DNA-coated MOFs may
one day become the standard way to carry life-saving protein drugs into our
cells.